Human cytomegalovirus (HCMV) is an important human pathogen and a major opportunist which emerges to cause disease in immuno-compromised individuals such as AIDS patients, neonates, and individuals who have been given immunosuppressive drugs as part of a transplantation regimen. In these individuals, the consequences of HCMV in acute or re-emerging infections can be dire, including retinitis, encephalitis, and pneumocystis, among other pathologies. Furthermore, in immuno-competent hosts, HCMV establishes a persistent lifelong infection through which it has been linked to a variety of inflammatory conditions including coronary artery occlusion following heart transplant and arthrectomy and restenosis following angioplasty. Additionally, following infection or reinfection of an immune-competent HCMV sero-negative woman during pregnancy, transmission of virus to the fetus may result in congenital infection of the child with severe physical and/or mental abnormalities resulting.
The genome (230 kb) of HCMV shares certain structural similarities with herpes simplex virus. In particular, HCMV includes a long and short unique region (UL and US, respectively), each of which is flanked by inverted repetitions. The entire HCMV genome has been sequenced (Chee, M. S., et al. (1990) Curr. Top. Microbiol. Immunol. 154:125-169) and appears to contain over 200 open reading frames.
One of these open reading frames is referred to as US28, which encodes a protein (also, “US28”) that acts as a functional receptor for certain human and viral chemokines (see, e.g., Gao & Murphy, 1994, J Biol Chem. 269:28539-42). Upon infection of a cell by CMV, US28 is expressed on the surface of the infected cell and becomes capable of responding to chemokines in the environment. Three other open reading frames called US27, UL33 and UL78 encode for proteins having homology to US28 as shown in Table 1 below.
TABLE 1Exemplary Viral Chemokine Elements and Immune-inhibitory GenesCMV ChemokineElements orGenBankImmune-Accessioninhibitory GenesNo.ReferenceUS27X17403Chee et al, 1990, Nature, 344:774US28L20501,Neote et al, 1993, Cell, 72:415-25AF073831-35UL33X53293Chee et al, 1990, Nature, 344:774UL78X17403Chee et al, 1990, Nature, 344:774
Chemokine receptors such as US28 generally are G protein coupled receptors. Structurally these receptors have seven transmembrane segments that loop in and out of the cell membrane, as well as an intracellular tail that is coupled to a G protein signal transducing molecular complex.
The chemokines themselves constitute a subgroup of a larger class of signaling proteins and have the ability, among other things, to promote cellular migration (Zlotnik et al. (1999) Crit. Rev. Immunol. 19:1-47). The chemokines generally are divided into four groups based upon the arrangement of certain cysteine residues within the protein that can form disulfide bonds. One class of chemokines are the beta chemokines which are characterized by having two adjacent cysteines; this structure is referred to in shorthand form simply as CC. The beta chemokines are involved in attraction of monocytes and leukocytes. The alpha chemokines in contrast have a single amino acid separating the two cysteine residues, and thus their structure is designated as CXC. These chemokines are primarily involved in attracting polymorphonuclear cells. The fractalkines constitute a third class of chemokines and tend to be cell bound molecules. The two cysteines in this class are separated by three amino acid residues, a structure designated as CX3C. This class of chemokines are expressed at high levels in the brain; some evidence indicates that the fractalkines are involved in neuron-glial cell interactions (see, e.g., Harrison, et al. (1998) Proc. Natl. Acad. Sci. U.S.A. 95:10896-10901; and Nishiyori, A. et al. (1998) FEBS Lett. 429:167-172). The structure of the final class of chemokines is simply referred to as C, because these chemokines contain only a single cysteine involved in a disulfide bond. The chemokine receptors have varying specificity for the different classes of chemokines. Some chemokine receptors can bind chemokines from different classes.
The US28 receptor of HCMV is characterized in part by its very strong affinity for fractalkine. It is unclear, however, whether this interaction is involved in the dissemination of the virus from the primary site of infection to other secondary sites. In fact, there is very little known concerning the mechanism by which HCMV is disseminated. Insight into this issue has been hampered primarily by the lack of assay systems, particularly in vivo assay systems, which are good models of viral dissemination in humans. Similarly, the lack of appropriate assay systems has hindered identification of inhibitors effective in reducing the spread of CMV infection.